中国组织工程研究

中国组织工程研究 ›› 2022, Vol. 26 ›› Issue (10): 1530-1536.doi: 10.12307/2022.199

• 组织工程口腔材料 tissue-engineered oral materials • 上一篇    下一篇

纳米改性聚己内酯微球的构建及对牙髓细胞的生物学效应

李  璇，孙一民，李龙飙，王振铭，杨  静，汪成林，叶  玲   

  1. 四川大学华西口腔医学院，四川省成都市   610041
  • 收稿日期:2020-10-12 修回日期:2020-10-14 接受日期:2020-11-28 出版日期:2022-04-08 发布日期:2021-10-25
  • 通讯作者: 叶玲，博士，教授，四川大学华西口腔医学院，四川省成都市 610041
  • 作者简介:李璇，女，1995年生，山东省潍坊市人，汉族，四川大学华西口腔医学院在读博士，主要从事牙髓及骨组织工程方面的研究
  • 基金资助:
    四川省科技厅应用基础研究项目(2019YFS0035)，项目负责人：叶玲；国家自然科学基金杰出青年科学基金项目(81825005)，项目负责人：叶玲

Manufacturing of nano-modified polycaprolactone microspheres and its biological effects in dental pulp cells

Li Xuan, Sun Yimin, Li Longbiao, Wang Zhenming, Yang Jing, Wang Chenglin, Ye Ling   

  1. West China School of Stomatology, Sichuan University, Chengdu 610041, Sichuan Province, China
  • Received:2020-10-12 Revised:2020-10-14 Accepted:2020-11-28 Online:2022-04-08 Published:2021-10-25
  • Contact: Ye Ling, MD, Professor, West China School of Stomatology, Sichuan University, Chengdu 610041, Sichuan Province, China
  • About author:Li Xuan, Doctoral candidate, West China School of Stomatology, Sichuan University, Chengdu 610041, Sichuan Province, China
  • Supported by:
    the Foundation of Basic Research for Application from the Department of Science and Technology, Sichuan Province, No.2019YFS0035 (to YL); the National Science Foundation for Distinguished Young Scholars of China, No.81825005 (to YL) 

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摘要： 文题释义：
聚己内酯-多巴胺-纳米羟基磷灰石微球：在聚己内酯多孔微球表面修饰多巴胺，以增加其亲水性和结晶位点，随后在模拟体液原位形成纳米羟基磷灰石，从而得到表面改性的聚己内酯纳米微球。
支架材料：组织工程必备的3个条件之一，在组织工程中支架可作为细胞运输的载体，为细胞的定植、黏附、增殖和分化提供三维空间环境，并在细胞外基质的形成过程中被逐步降解。

背景：聚己内酯具有良好的热稳定性、生物相容性、降解速率可调等优点，有作为组织工程支架应用于牙髓组织工程的潜能，但其亲水性和生物活性较差。
目的：制备聚多巴胺及纳米羟基磷灰石改性的多孔聚己内酯(polycaprolactone-polydopamine-nano-hydroxyapatite，PCL-PDA-nHA)微球，探索其物理性能及对牙髓细胞增殖和矿化的影响。
方法：采用双乳化法制备多孔聚己内酯微球，通过聚多巴胺进行表面改性，得到PCL-PDA微球，提高其亲水性和结晶能力；通过模拟体液在PCL-PDA微球表面原位形成纳米羟基磷灰石涂层(分反应7 d组与反应14 d组)，得到PCL-PDA-nHA微球。通过扫描电镜、X射线光电子能谱技术和蛋白吸附实验检测各组微球的理化性质，通过溶血实验、凝血因子激活实验和血小板凝集实验检验各组微球的血液相容性。将4组微球分别与人牙髓细胞共培养，CCK-8法检测细胞增殖，碱性磷酸酶、茜素红与天狼星红染色分析微球的矿化诱导能力。

结果与结论：①扫描电镜显示，4组微球为均匀疏松的多孔结构，微球直径无明显差异，微球孔隙率为87.4%，孔隙直径在20-50 μm之间；X射线光电子能谱显示，聚多巴胺和纳米羟基磷灰石有效修饰到聚己内酯表面；蛋白吸附实验显示，改性后的材料可增强血清白蛋白的吸附能力；②溶血实验显示，4组微球的溶血率在1%以下，不会造成明显的溶血；凝血因子激活实验显示，各组改性聚己内酯微球对凝血影响较小；扫描电镜显示，各组微球表面无明显的血小板聚集；③CCK-8检测显示，各组改性微球表面的细胞增殖均快于聚己内酯微球；④碱性磷酸酶、茜素红与天狼星红染色显示，各组改性微球的矿化诱导能力均强于聚己内酯微球，其中PCL-PDA-nHA-14 d微球与  PCL-PDA-nHA-7 d微球的矿化诱导能力强于PCL-PDA微球；⑤结果表明，PCL-PDA-nHA纳米微球的血液相容性良好，可促进人牙髓细胞的增殖和矿化。

https://orcid.org/0000-0003-2452-3335 (李璇) 


关键词: 材料, 聚多巴胺, 聚己内酯, 羟基磷灰石, 多孔材料, 牙髓组织工程, 支架材料, 人牙髓细胞, 细胞增殖, 促矿化

Abstract: BACKGROUND: Polycaprolactone based scaffolds have potential in dental pulp regeneration since its good thermal stability, excellent biocompatibility and tunable degradation rate. However, its hydrophilicity and bioactivity are poor.    
OBJECTIVE: To prepare microspheres with polycaprolactone-polydopamine-nano-hydroxyapatite (PCL-PDA-nHA), and explore its physical properties and the effects on the proliferation and mineralization of dental pulp cells.
METHODS: The PCL microspheres were prepared by double emulsification method. The hydrophilicity and crystallinity were improved by the surface modification of dopamine and the PCL-PDA particles were obtained. Nano-hydroxyapatite coating was formed in situ on the surface of PCL-PDA microspheres by simulating body fluids (reaction 7-day group and reaction 14-day group) to obtain PCL-PDA-nHA microspheres. The physical and chemical properties of each group of microspheres were detected by scanning electron microscopy, X-ray photoelectron spectroscopy and protein adsorption experiment. Hemolysis test, coagulation factor activation test and platelet agglutination test were used to test the hemocompatibility of each group of microspheres. Four groups of microspheres were co-cultured with human dental pulp cells. Cell proliferation was detected by CCK-8 assay. The mineralization induction ability of the microspheres was analyzed using alkaline phosphatase, alizarin red and sirius red staining.
RESULTS AND CONCLUSION: (1) Scanning electron microscopy showed that the four groups of microspheres were uniform and porous structure, and there was no significant difference in the diameter of microspheres; the porosity was 87.4% with pore diameter ranging from 20 μm to 50 μm. X-ray photoelectron spectroscopy showed that polydopamine and hydroxyapatite particles were successfully decorated on the microsphere surface. Protein adsorption experiment showed that the functional microspheres could enhance the adsorption of serum albumin. (2) Hemolysis experiment showed that the hemolysis rate of microspheres was below 1%, without obvious hemolysis. The coagulation factor activation experiment showed that each group of modified polycaprolactone microspheres had little effect on blood coagulation. Scanning electron microscopy showed that there was no obvious platelet aggregation on the surface of each group of microspheres. (3) CCK-8 assay demonstrated that cell proliferation on the surface of each group of modified microspheres was faster than that of PCL microspheres. (4) Alkaline phosphatase, alizarin red and sirius red staining exhibited that the mineralization induction ability of each group of modified microspheres was stronger than that of PCL microspheres, and the mineralization induction ability of PCL-PDA-nHA-14 day microspheres and PCL-PDA-nHA-7 day microspheres was stronger than that of PCL-PDA microspheres. (5) The results suggested that PCL-PDA-nHA nanospheres had good hemocompatibility, and could promote the proliferation and mineralization of human dental pulp cells. 

Key words: material, polydopamine, polycaprolactone, hydroxyapatite, porous materials, dental pulp tissue engineering, scaffold material, human pulp cells, cell proliferation, mineralization

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